South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, PR China; College of Aqua-life Science and Technology, Shanghai Ocean University, Shanghai, PR China; Key Laboratory of South China Sea Fishery Resources Exploitation & Utilization, Ministry of Agriculture, Guangzhou, PR China.
South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, PR China; Key Laboratory of South China Sea Fishery Resources Exploitation & Utilization, Ministry of Agriculture, Guangzhou, PR China; Key Laboratory of Fishery Ecology and Environment, Guangdong Province, PR China.
Fish Shellfish Immunol. 2023 Oct;141:109065. doi: 10.1016/j.fsi.2023.109065. Epub 2023 Sep 7.
Thymosin beta-4 (Tβ4) is a ubiquitous protein with multiple and diverse intracellular and extracellular functions in vertebrates, which play fundamental roles in innate immune against pathogens and wound healing. In this study, the full-length cDNA of Tβ4 was cloned from Penaeus monodon (designated as PmTβ4), using the technology of rapid amplification of cDNA ends (RACE). The cDNA of PmTβ4 was 1361 bp with an open reading frame (ORF) of 501 bp, which encoding a polypeptide of 166 amino acid. The Quantitative Real-time PCR (qRT-PCR) analysis results showed that PmTβ4 was ubiquitously expressed in all the tested shrimp tissues, with the highest expression level was detected in the hemolymph, while the lowest expression level in the muscle. The expression level of PmTβ4 was significantly up-regulated in hepatopancreas after challenged by Vibrio parahaemolyticus, Vibrio harveyi and Staphylococcus aureus. In vitro antimicrobial test showed that the recombinant protein of PmTβ4 (rPmTβ4) had broad-spectrum of antimicrobial activity, which could inhibit both the growth of gram-negative bacteria and gram-positive bacteria, including Vibrio vulnificus, V. parahaemolyticus, Streptococcus agalactiae, S. aureus and Aeromonas hydrophila. Moreover, rPmTβ4 had a certain binding ability to different bacteria, and this binding ability exhibits a strong dose-dependent effect. In vivo, PmTβ4 could facilitate external bacterial clearance in shrimp, and have beneficial to shrimp survival post V. parahaemolyticus infection. Furthermore, wound-healing assay was carried out to study the role of PmTβ4 in the process of wound healing. The results showed that the PmTβ4 expression was significantly up-regulated by injury treatment, and exerted positive effects to promote wound healing. In addition, PmTβ4 can significantly increase the expression level of superoxide dismutase (SOD) and Catalase (CAT) after injury treatment in shrimp, which would involve in scavenging reactive oxygen species (ROS) caused by the wound. In conclusion, these results indicated that PmTβ4 may play important roles in antibacterial immunity and wound healing in Penaeus monodon.
胸腺素β-4（Tβ4）是一种普遍存在的蛋白质，在脊椎动物中有多种不同的细胞内和细胞外功能，在先天免疫对抗病原体和伤口愈合中发挥着重要作用。在这项研究中，使用快速扩增 cDNA 末端（RACE）技术，从斑节对虾（命名为 PmTβ4）中克隆了 Tβ4 的全长 cDNA。PmTβ4 的 cDNA 长 1361bp，开放阅读框（ORF）长 501bp，编码 166 个氨基酸的多肽。定量实时 PCR（qRT-PCR）分析结果表明，PmTβ4 在所有测试的虾组织中广泛表达，在血淋巴中的表达水平最高，而在肌肉中的表达水平最低。在受到副溶血弧菌、哈维弧菌和金黄色葡萄球菌的刺激后，肝胰腺中的 PmTβ4 表达水平显著上调。体外抗菌试验表明，PmTβ4 的重组蛋白（rPmTβ4）具有广谱抗菌活性，能抑制革兰氏阴性菌和革兰氏阳性菌的生长，包括创伤弧菌、副溶血弧菌、无乳链球菌、金黄色葡萄球菌和嗜水气单胞菌。此外，rPmTβ4 与不同细菌具有一定的结合能力，这种结合能力呈强剂量依赖性。在体内，PmTβ4 可以促进虾体内外细菌的清除，有助于虾在感染副溶血弧菌后的存活。此外，进行了伤口愈合试验以研究 PmTβ4 在伤口愈合过程中的作用。结果表明，损伤处理后 PmTβ4 的表达明显上调，并对促进伤口愈合产生积极影响。此外，损伤处理后，PmTβ4 可显著提高虾中超氧化物歧化酶（SOD）和过氧化氢酶（CAT）的表达水平，从而清除伤口引起的活性氧（ROS）。综上所述，这些结果表明，PmTβ4 在斑节对虾的抗菌免疫和伤口愈合中可能发挥重要作用。
